Phosphodiesterase-5 (PDE-5) inhibitor, sildenafil (Viagra), which is known to enhance erectile function in men, induces powerful cardioprotective effect against ischemia-reperfusion injury (I/R) in heart. We propose to perform further in depth studies on the mechanisms by which sildenafil and other clinically used PDE-5 inhibitors trigger signaling pathways that lead to cardioprotection. Hypothesis 1: Novel class of PDE-5 inhibitors trigger protection against I/R in heart and myocytes. We will study the effect of short acting (sildenafil, verdenafil) or long-acting (tadalafil) PDE-5 inhibitors on protective effect against I/R and apoptosis. Hypothesis 2: Release of preconditioning mediators, including adenosine, bradykinin and nitric oxide mediate cardioprotection induced by PDE-5 inhibitors. Using pharmacological inhibitors and gone knockout mice of adenosine A1, bradykinin B2 receptors, eNOS, nNOS and iNOS. we will determine the direct role of these mediators in cardioprotection by PDE-5 inhibitors. Hypothesis 3: PDE-5 inhibitors selectively activate PKC isoforms that play an essential role in the gene transcription of eNOS/iNOS/nNOS to induce cardioprotective effect. We will study translocation of PKC isozymes after treatment with PDE-5 inhibitors. Using selective peptide blockers of PKC isozymes, we will determine their role in gone transcription of eNOS, iNOS or nNOS. Hypothesis 4:PDE-5 inhibitors stimulate guanylate cyclase and elevate cGMP causing activation of protein kinase G (PKG) as a prelude to the opening of mitoKATP channels. Using adenoviral gene transfer of splice variants of PKG or their null mutants, we will determine their role in protection against I/R in cardiomyocytes and intact heart. Hypothesis 5:PDE-5 inhibitors protect against I/R by overexpression of HSPs through activation of transcription factor, HSF-1. Using siRNAs, we will determine the cause and effect relationship of HSF-1 in cardioprotection by PDE-5 inhibitors in mice. These studies, first of its kind, will provide tremendous amount of new information on protective effect of PDE-5 inhibitors against I/R injury. It is expected that the studies will have enormous impact on bringing the long-studied phenomenon of ischemic and pharmacologic preconditioning to the clinical forefront. [unreadable] [unreadable]